Cloning, purification and biochemical characterization of recombinant Cathepsin L from Takifugu rubripes and its role in taste formation

  • Qilong Tang
  • Wenli Wang
  • Lujia Zhang
  • Yuan LiuEmail author
Original Paper


In recent years Takifugu rubripes has been increasingly popular among people base on its good taste. Cathepsin L was related to the formation of amino acids and small peptides. This study described the biochemical characterization of Cathepsin L and its role in the taste formation of Takifugu rubripes. Cathepsin L gene from Takifugu rubripes tissues was cloned successfully, and Cathepsin L with molecular masses of 46 kDa was isolated and enzymatically characterized. This enzyme reached its highest activity at 40 °C and pH 5.5. Cu2+ and Mn2+ ions reduced the activity of Cathepsin L, but Fe3+ and Ca2+ ions prominently increased its activity when the final concentrations of metal ions were 1 mM and 5 mM, respectively. The addition of phenylmethanesulfonyl fluoride gradually decreased the enzyme activity over 0.5 mM. 200–3000 KDa fraction from Takifugu rubripes muscle after Cathepsin L hydrolysis showed the stronger kokumi and umami taste, especially kokumi. In this study, the recombinant Cathepsin L from Takifugu rubripes could play a role in flavor formation and be applied in flavor studies through the degradation of animal proteins.


Takifugu rubripes Cathepsin L Biochemical characterization Flavor peptides 



This work was funded by The National Key R&D Program of China (2016YFD0400803, 2016YFD0401501) and National Natural Science Foundation of China (Grant Nos. 31622042, 31371790, 31271900).

Compliance with ethical standards

Conflict of interest

The authors declared that they had no conflict of interest.

Supplementary material

11694_2019_122_MOESM1_ESM.docx (50 kb)
Supplementary material 1 (DOCX 49 kb)


  1. 1.
    A.J. Barrett, Controversies in Neuro-urology (Wiley, New Jersey, 2008), pp. 3–16Google Scholar
  2. 2.
    J. Liu, G.K. Sukhova, J.T. Yang, J.S. Sun, L.K. Ma, A. Ren, W.H. Xu, H.X. Fu, G.M. Dolganov, C.C. Hu, P. Libby, G.P. Shi, Atherosclerosis 184(2), 302–311 (2006)CrossRefGoogle Scholar
  3. 3.
    I. Whang, M. De Zoysa, C. Nikapitiya, Y. Lee, Y. Kim, S. Lee, C. Oh, S.J. Jung, M.J. Oh, C.Y. Choi, S.Y. Yeo, B.S. Kim, S.J. Kim, J. Lee, Fish Shellfish Immunol. 30(3), 763–772 (2011)CrossRefGoogle Scholar
  4. 4.
    Q.H. Li, J.Q. Ao, Y.N. Mu, Z.J. Yang, T. Li, X. Zhang, X.H. Chen, Fish Shellfish Immunol. 47(2), 743 (2015)CrossRefGoogle Scholar
  5. 5.
    J.Z. Liang, Y.Z. Rao, Z.R. Lun, T.B. Yang, Fish Physiol. Biochem. 38(6), 1795–1806 (2012)CrossRefGoogle Scholar
  6. 6.
    Y.X. Sun, L. Tang, P. Wang, M.N. Abbas, J.W. Tian, B.J. Zhu, C.L. Liu, Dev. Comp. Immunol. 78, 114–123 (2018)CrossRefGoogle Scholar
  7. 7.
    L.S. Dai, S.H. Chu, X.M. Yu, Y.Y. Li, Fish Shellfish Immunol. 71, 246–254 (2017)CrossRefGoogle Scholar
  8. 8.
    W.J. Wang, L. Long, L. Wang, C.H. Tan, X.F. Fei, L.S. Chen, Q. Huang, Z.Q. Liang, Cancer Lett. 371(2), 274–284 (2016)CrossRefGoogle Scholar
  9. 9.
    K.L. Mattock, P.J. Gough, J. Humphries, K. Burnand, L. Patel, K.E. Suckling, F. Cuello, C. Watts, M. Gautel, M. Avkiran, A. Smith, Atherosclerosis 208(1), 83–89 (2010)CrossRefGoogle Scholar
  10. 10.
    Y. Zhong, J. Zhao, Y.J. Gu, Y.F. Zhao, Y.W. Zhou, G.X. Fu, Arch. Gerontol. Geriat. 61(2), 285–288 (2015)CrossRefGoogle Scholar
  11. 11.
    L.A. Pennacchio, Science 2(5256), 1731–1734 (1996)CrossRefGoogle Scholar
  12. 12.
    P. Taupin, J. Ray, W.H. Fischer, S.T. Suhr, K. Hakansson, A. Grubb, F.H. Gage, Neuron 28(2), 385–397 (2000)CrossRefGoogle Scholar
  13. 13.
    J. Palomino, G. Herrera, J. Torres-Fuentes, P. Dettleff, A. Patel, V. Martinez, Anim. Reprod. Sci. 180, 23–29 (2017)CrossRefGoogle Scholar
  14. 14.
    F. Toldra, M.N. Flores, Crit. Rev. Food Sci. 38(4), 331 (1998)CrossRefGoogle Scholar
  15. 15.
    R. Cheret, C. Delbarre-Ladrat, M. de Lamballerie-Anton, V. Verrez-Bagnis, Food Chem. 101(4), 1474–1479 (2007)CrossRefGoogle Scholar
  16. 16.
    L.S. Song, B.Z. Liu, J.H. Xiang, P.Y. Qian, Mar. Biotechnol. 3(4), 398–406 (2001)CrossRefGoogle Scholar
  17. 17.
    M.X. Zhang, X.C. Wang, Y. Liu, X.L. Xu, G.H. Zhou, Food Chem. 135(3), 1463–1470 (2012)CrossRefGoogle Scholar
  18. 18.
    M.X. Zhang, X.C. Wang, Y. Liu, X.L. Xu, G.H. Zhou, Sensors 12(9), 12562–12571 (2012)CrossRefGoogle Scholar
  19. 19.
    M. Wang, X.Y. Zhang, R.R. Guo, Z.P. Cai, X.C. Hu, H. Chen, S. Wei, J. Voglmeir, L. Liu, Carbohyd. Res. 457, 1–7 (2018)CrossRefGoogle Scholar
  20. 20.
    M. Flores, M.C. Aristoy, T. Antequera, J.M. Barat, F. Toldra, Meat Sci. 82(2), 241–246 (2009)CrossRefGoogle Scholar
  21. 21.
    X.Q. Yu, L.J. Zhang, X.D. Miao, Y.Y. Li, Y. Liu, Food Chem. 221, 599–605 (2017)CrossRefGoogle Scholar
  22. 22.
    X. Niu, M.J. Guiltinan, Nucleic Acids Res. 22(23), 4969–4978 (1994)CrossRefGoogle Scholar
  23. 23.
    A.K. Mohanty, M.C. Wiener, Protein Expr. Purif. 33(2), 311–325 (2004)CrossRefGoogle Scholar
  24. 24.
    R.M. Nepal, S. Mampe, B. Shaffer, A.H. Erichson, P. Bryant, Int. Immunol. 18(6), 931–939 (2006)CrossRefGoogle Scholar
  25. 25.
    F. Lecaille, E. Authie, T. Moreau, C. Serveau, F. Gauthier, G. Lalmanach, Eur. J. Biochem. 268(9), 2733 (2001)CrossRefGoogle Scholar
  26. 26.
    K. Schilling, S. Pietschmann, M. Fehn, I. Wenz, B. Wiederanders, Biol. Chem. 382(5), 859 (2001)CrossRefGoogle Scholar
  27. 27.
    M.C. Hughes, E.E. Neill, P.L.H. McSweeney, Food Chem. 64(4), 525–530 (1999)CrossRefGoogle Scholar
  28. 28.
    F. Shahidi, Y.V.A.J. Kamil, Trends Food Sci. Tech. 12(12), 435–464 (2001)CrossRefGoogle Scholar
  29. 29.
    D.P. Bown, H.S. Wilkinson, M.A. Jongsma, J.A. Gatehouse, Insect Biochem. Mol. 34(4), 305–320 (2004)CrossRefGoogle Scholar
  30. 30.
    Y.Y. Tian, E. Umezawa, R. Duan, K. Konno, Fisheries Sci. 76(2), 365–373 (2010)CrossRefGoogle Scholar
  31. 31.
    T. Yoonuan, S. Nuamtanong, P. Dekumyoy, O. Phuphisut, P. Adisakwattana, Parasitol. Res. 115(12), 4457–4470 (2016)CrossRefGoogle Scholar
  32. 32.
    N. Rotzoll, A. Dunkel, T. Hofmann, J. Agr. Food Chem. 53(10), 4149–4156 (2005)CrossRefGoogle Scholar
  33. 33.
    O. Frank, H. Ottinger, T. Hofmann, J. Agr. Food Chem. 49(1), 231–238 (2001)CrossRefGoogle Scholar
  34. 34.
    M.R. Kim, Y. Kawamura, K.M. Kim, H. Cherl, M.R. Lee, J. Agr. Food Chem. 56(14), 5852–5858 (2008)CrossRefGoogle Scholar

Copyright information

© Springer Science+Business Media, LLC, part of Springer Nature 2019

Authors and Affiliations

  1. 1.College of Food Science and TechnologyShanghai Ocean UniversityShanghaiChina
  2. 2.College of Life SciencesEast China Normal UniversityShanghaiChina
  3. 3.College of Agriculture and BiologyShanghai Jiao Tong UniversityShanghaiChina

Personalised recommendations